It is proposed to continue to complete the purification of the SV40 tumor antigen to homogeneity from both transformed and lytically infected cells. We wish to ascertain, as accurately as possible, the molecular weights of these proteins. We would also hope to be able to develop a radioimmunoassay for T antigen. Attempts will be made to ascertain the amino terminal sequence of T antigen from both transformed and lytically infected cells in an effort to discern where in the early cistron the minimum 5' translational start point is located. We also plan to locate with precision the binding sites for SV40 T antigen which we have recently identified on the SV40 genome. We would like to determine the nucleotide sequence of these binding regions and to ascertain which residues within these regions constitute contact points for the protein. An attempt will be made to microinject SV40 T antigen from both transformed and lytically infected cells into permissive and non-permissive cells in an effort to ask whether the protein is capable of inducing rounds of viral DNA replication and/or initiating and/or establishing an abortive state of neoplastic transformation. Lastly, an effort will be made to ascertain whether binding specific individual loci on the viral genome is of physiological consequence and also whether the antigen has site-specific nuclease or nucleotide or deoxynucleotide polymerizing or certain other enzymatic activities associated with polynucleotide replication.